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. Author manuscript; available in PMC: 2015 Dec 1.
Published in final edited form as: Mol Microbiol. 2014 Sep 17;94(6):1227–1241. doi: 10.1111/mmi.12780

Figure 1.

Figure 1

Genetic disruption of the housekeeping sortase srtA in A. oris. (A) Presented is the non-replicative vector pCWU2 used to generate unmarked, in-frame gene deletions in A. oris. The vector carries 1-kb flanking regions of srtA, the kanamycin resistance cassette (kanR), and galK. Brackets indicate regions amplified by a primer pair P1 and P2. (B) Shown is the schematic of srtA allelic replacement using pCWU2-ΔsrtA. Integration of pCWU2-ΔsrtA into the A. oris ΔgalK mutant CW1 generates a srtA/ΔsrtA merodiploid strain (Int). Excision of srtA from the Int chromosome is permitted in the presence of a plasmid expressing SrtA under the control of a tetracycline-inducible promoter, resulting srtA mutant and wild-type alleles. (C) Excision of srtA from the chromosome of mutant candidates is confirmed by PCR-amplification using the primer pairs P1 and P2.